Protein Accumulation Disorders
Many neurological diseases are associated with protein accumulation and aggregation in the brain. For example, Alzheimer's Disease involves accumulation of the Aβ protein, Parkinson's Disease involves accumulation of α-synuclein, Huntington's Disease involves aggregation of mutated huntingtin proteins, and amyotrophic lateral sclerosis involves accumulation of mutated superoxide dismutase-1 proteins. Some chronic psychiatric disorders, such as schizophrenia, bipolar disorder, and recurrent major depression, have also been associated with protein aggregation. The reduction of protein accumulation events is important for slowing the progression of these diseases and disorders. Studies indicate that protein degradation processes that clear these aggregated proteins could provide treatment for some or all of these diseases and disorders.
Alzheimer's Disease
The age-related neurodegenerative disorder Alzheimer's Disease (Alzheimer's) involves the accumulation of oligomeric species, protein aggregation, and altered brain function. One of the major hallmarks of Alzheimer's is the plaque deposits consisting primarily of amyloid fibrils formed by the amyloid beta peptide Aβ1-42 as well as the buildup of soluble oligomers of this peptide. Mutations associated with familial Alzheimer's, including mutations in the amyloid precursor protein (APP), strongly implicate Aβ1-42 as a causative factor since the mutations increase the relative amount of this Aβ peptide. Increased Aβ is one of the earliest events in Alzheimer's, and, besides extracellular accumulation, Aβ oligomerization also occurs intraneuronally. Aβ oligomers disrupt synaptic plasticity, impair synaptic responses and memory, and cause cytotoxicity, as well as produce synaptic deterioration. Aβ oligomers, especially trimers and multiples of trimeric species, are particularly stable.
There are no current treatments to reduce the abnormal protein accumulation events in Alzheimer's. Only two classes of drugs are approved for treating Alzheimer's, acetyl-cholinesterase inhibitors and N-methyl-D-aspartic acid (NMDA) receptor antagonists. Both types of drugs only affect the symptoms of Alzheimer's. Acetyl-cholinesterase inhibitors are for mild to moderate Alzheimer's and have modest effects in a small percentage of patients who take the drug, and are typically ineffective after 6-12 months of use. The NMDA receptor antagonist that is available treats the secondary pathology but not the protein accumulation in mild to severe Alzheimer's.
Parkinson's Disease
Parkinson's Disease (Parkinson's) is a motor system disorder which is associated with the loss of dopamine-producing brain cells. Dopamine is necessary for coordinated muscle function and movement. Dopamine is normally produced by certain nerve cells (neurons) in the substantia nigra region of the brain; however, Parkinson's patients experience a loss of these neurons which leads to impaired movement. This loss of neurons is associated with the accumulation of alpha synuclein, a protein that is mutated and/or misfolded in Parkinson's and other diseases. The alpha synuclein forms aggregates that accumulate in Lewy bodies, and which are seen in the brains of patients who have died from Parkinson's.
Huntington's Disease
The neurodegenerative disorder Huntington's Disease (Huntington's) is caused by a trinucleotide repeat expansion in the huntingtin gene which codes for huntingtin protein, “Htt.” People who have Huntington's Disease have more C-A-G codons on their huntingtin gene which results in Htts that are “altered” or abnormal in that they have an excess number of glutamines. As a result of the excess glutamines, these altered Htts form protein aggregates which can interfere with nerve cell function.
Amyotrophic Lateral Sclerosis
Amyotrophic lateral sclerosis (ALS) is a progressive neurodegenerative disease that attacks nerve cells in the brain and the spinal cord. Neuronal cell death has been linked to the presence of aggregates of mutant superoxide dismutase-1 (SOD1) protein. Mutant SOD1 accumulates to form high molecular weight amorphous aggregates which can interact with other proteins. When these mutant SOD1 proteins accumulate and form aggregates in a neuronal cell, the cell almost always dies.
Chronic Psychiatric Disorders
Studies on patients with phenotypes of affective disorders or schizophrenia have shown that significant fractions of the disrupted-in-schizophrenia (DISC1) protein form aggregates identified as cold Sarkosyl-insoluble protein aggregates. These aggregates do not bind with nuclear distribution element 1 (NDEL1), a key DISC1 ligand, demonstrating a loss of function phenotype. Specifically, in human neuroblastoma cells the aggregates are expression-dependent, detergent-resistant and do not interact with endogenous NDEL1. Escherichia coli expresses recombinant (r) NDEL1 which selectively binds to an octamer of an rDISC1 fragment. The rNDEL1 does not bind with dimers or with high molecular weight multimers. Thus, for molecular interaction of DISC1 with NDEL1, an optimum oligomerization exists which is exceeded by aggregation of DISC1. The absence of oligomer-dependent interactions of DISC1 is associated with sporadic mental disease of mixed phenotypes.
Lysosomal Modulation
Lysosomes represent a major degradation pathway necessary for cells to maintain normal protein turnover. The lysosomal pathway is believed to contribute to the clearance of Aβ oligomers, and reduced efficiency in this pathway can have detrimental effects. Administering high dosages of lysosomal inhibitors into brain ventricles of rats, systemically in mice, and to hippocampal slice cultures has been shown to result in the buildup of Aβ species. In addition to Aβ oligomers forming intraneuronally, released Aβ peptide can also be taken up by Alzheimer's-vulnerable neurons causing Aβ accumulation in lysosomes, lysosomal disruption, and the further production of amyloidogenic species. Disturbances in lysosomes have a marked prevalence in Alzheimer's-vulnerable brain regions and are particularly evident in the aged brain, familial Alzheimer's, and related transgenic mouse models.
Lysosomes play an important role in normal protein turnover as well as in clearing and preventing the buildup of misfolded or damaged proteins. Degradation of long-lived proteins and clearance of toxic accumulation events occur in large part via lysosomes, as well as via lysosomes fused with autophagolysosomes of the autophagy pathway. For example, cathepsin is a lysosomal enzyme that cleaves Aβ1-42 peptide to smaller non-pathogenic species. This cleavage reduces the amount of Aβ1-42 in the brain and contributes to the restoration of synaptic integrity. Overall compromise of lysosomes leads to the accumulation of Aβ and other proteins, and several reports indicate that enhancement of lysosomal function is a plausible strategy to reduce protein accumulation events in several age-related neurodegenerative disorders.
Lysosomes and autophagolysosomes are thought to be activated for the clearance of toxic material. As toxic protein species accumulate, autophagy-lysosomal pathways show clear signs of activation, perhaps as distinct compensatory responses. Responses include autophagic vacuoles and enhanced levels of lysosomal hydrolases including cathepsins B and D. The cathepsin family of lysosomal proteases appears to be particularly responsive to accumulating proteins in neurons. Protein accumulation stress, including that produced by the aggregation-prone Aβ1-42 peptide, up-regulates the message, protein, and activity levels of cathepsins. Such responses may keep protein accumulation events partially in check and account for the gradual nature of the associated pathology that can extend over many years in Alzheimer's patients. Cathepsin B, in particular, is a lysosomal enzyme found to reduce Aβ1-42 deposition by cleaving the peptide into non-pathogenic species.
Many studies indicate that induction of protein degradation processes occurs as an attempt to clear Aβ and tau species in Alzheimer's, α-synuclein in Parkinson's Disease, and mutant huntingtin in Huntington's Disease. Lysosomal responses are also common among lysosomal storage disorders (LSDs). Mutations that cause specific enzyme deficiencies account for most of these diseases. Metabolically mutated animals comparable to Niemann-Pick Disease and Gaucher's Disease exhibit elevated activities of several lysosomal hydrolases during cellular accumulation events. In addition, mannose 6-phosphorylated glycoproteins and hydrolases were found elevated in the brain in juvenile neuronal ceroid lipofuscinoses. The increased lysosomal enzyme activities are a likely indicator of cellular responses aimed at compensating for accumulating material. It is of interest that caloric restriction enhances protein clearance processes through the increased expression of lysosomal enzymes, and the same treatment improves brain function in a LSD model.
Protein accumulation has been shown to increase the expression of cathepsins, and this response allows for more efficient protein clearance. Such compensatory responses may delay overt neurodegeneration and account for the gradual nature of protein accumulation pathology that can extend over months or years in model systems and years or decades in Alzheimer's. The compensatory response is enhanced by modest levels of certain hydrolase inhibitors, including the cathepsin B and L inhibitor Z-Phe-Ala-diazomethylketone (PADK), making up a class of compounds deemed lysosomal modulators. The induced enhancement of lysosomal capacity has been found to far exceed the mild inhibitory action of PADK, thereby promoting the clearance of PHF-tau and other dementia-related proteins, re-establishing microtubule integrity and microtubule-based transport, and restoring synaptic markers in a brain slice model of protein accumulation pathology.
Drugs that increase lysosomal capacity promote clearance of protein species related to Aβ and tau that accumulate in Alzheimer's Disease and are valuable for treating accumulation events in Parkinson's, Huntington's, and other diseases and disorders. Moreover, enhanced clearance is associated with the restoration of synaptic integrity which is vital for neuronal communication mechanisms underlying memory function as well as neuronal maintenance. Lysosomal modulation represents a unique pharmacological strategy against Alzheimer's. Lysosomal modulators have several advantages over prior art methods: (1) they are first-in-class drugs for treating neurodegenerative disorders either alone or in combination with current treatments; (2) they promote clearance of a broad array of potentially pathogenic proteins (as compared to the host of strategies being developed to reduce Aβ or tau accumulations exclusively); and (3) the drugs repair multiple types of synapses important for memory (as compared to current treatments that act on cholinergic synapses only).
At very high levels PADK can have the opposite effect—i.e. can shut down lysosomal function and cause neuronal compromise. Thus, there is a need for lysosomal modulators that are less toxic, which would provide a safety margin between therapeutic dose and the dose that causes adverse effects. Furthermore, PADK is a peptidic compound and is likely to be unable to be used in vivo due to being rapidly metabolized and unable to penetrate the central nervous system.
Accordingly, lysosomal modulators are provided that are non-peptidic, have increased efficacy, demonstrate a better safety profile, and have improved stability, aqueous solubility, and bioavailability.